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In vitro peptidoglycan synthesis assay with lipid II substrate
Jacob Biboy1, Nhat Khai Bui, Waldemar Vollmer
1Institute for Cell and Molecular Biosciences, The Centre for Bacterial Cell Biology, Newcastle University, Newcastle upon Tyne, UK.
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Bacterial cell wall peptidoglycan is synthesized from lipid II precursor by two reactions. Glycosyltransferases polymerize the glycan chains and transpeptidases form the peptide cross-links. The bifunctional class A penicillin-binding proteins catalyze both of these reactions. Here, we describe an in vitro peptidoglycan synthesis assay utilizing radiolabeled lipid II substrate to monitor simultaneously peptidoglycan glycosyltransferase and transpeptidase activities. The products of the reaction are separated by high-pressure liquid chromatography and quantified by flow-through scintillation counting.
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